The plant Wedelia biflora has been used in traditional medicine in India and Vietnam to treat various symptoms. However, the chemical composition of its flowers remains mostly unknown. Therefore, we now report the isolation and structural elucidation of six new phenolic glycosides {wedebicoside A - F (1-6)} and one new ceramide [wedebiceramide (9)], together with six known compounds, 1-O-(2',4'-diangeloyloxy-beta-D-fucopyranosyl)-6-hydroxythymol (7), 1-O-[2',4'-diangeloyloxy-3'-(3"-angeloyloxy-beta-D-fucopyranosyl)-beta-D-fucopyransyl]-6-hydroxythymol (8), anhydrosecoisolariciresinol (10), friedeline (11), epifriedelanol (12) and stigmasterol (13) from the flowers of Wedelia biflora. Their structures were established by 1D and 2D NMR spectroscopy, as well as by high resolution ESI-MS analysis and comparison with literature data. The cytotoxic activities against HeLa, MCF-7 and NCI-H460 were evaluated on some purified compounds at the concentration of 100 microg/mL. Compounds 1, 2, 3 and 5 showed strong cytotoxic activities against three surveyed cancer cell lines. Consequently, this study elucidated the phytochemical composition of W. biflora, as well as the potential use of some of the new compounds against some cancers.